The project is designed to determine the regulatory effects of vitamin D3 and its important metabolites (25(OH)D3,24,25 (OH)2D3,1,25(OH)2D3) upon parathyroid gland function. Initial studies indicate suppressive effects of 1,25(OH)2D3 upon levels of preproPTH mRNA in cultured bovine parathyroid cells. The effects of vitamin D3 compounds on PTH secretion rates have also been examined and found to parallel those observed on PTH mRNA levels. The most active compound 1,25(OH)2D3 caused a marked decrease in PTH secretion, while the other metabolites had little or no effect. The time course, dose response and reversibility of these effects on both PTH mRNA levels and secretion rates were very similar. In addition, the suppressive effects of 1,25(OH)2D3 approached those seen with high Ca++ (2.5mM) and could not be completely overcome by incubation in low calcium (0.5mM). Studies will be performed to evaluate the mechanism of the suppressive effects of vitamin D3 metabolites. The studies will explore whether vitamin D3 metabolites regulate biosynthesis at the level of transcription or alter the stability of pre-proPTH mRNA. In addition, we will explore the possibility that vitamin D3 compounds affect PTH synthesis and secretion by a mechanism involving changes in Ca++. Cytosolic Ca++ concentrations of treated and control cells will be measured using the fluorescent cytosolic Ca++ indicator Quin II. Comparisons of normal bovine parathyroid cells with human adenomas and hyperplastic parathyroids will be made to search for possible differences exhibited in these pathologic tissues in vitro. In addition, the in vivo effects of vitamin D3 deprivation and treatment upon parathyroid gland function will be measured through determinations of preproPTH mRNA levels in parathyroids removed from D-deprived, normal and D-treated animals. Demonstration by these studies of suppressive effects of vitamin D3 compounds used in vivo may prove in the future to be of therapeutic benefit to patients exhibiting excessive parathyroid function in whom surgery is considered undesirable.